Fractionated Dosing Improves Preclinical Therapeutic Index of Pyrrolobenzodiazepine- Containing Antibody Drug Conjugates Mary Jane Masson Hinrichs1, Pauline M
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Published OnlineFirst June 19, 2017; DOI: 10.1158/1078-0432.CCR-17-0219 Cancer Therapy: Preclinical Clinical Cancer Research Fractionated Dosing Improves Preclinical Therapeutic Index of Pyrrolobenzodiazepine- Containing Antibody Drug Conjugates Mary Jane Masson Hinrichs1, Pauline M. Ryan1, Bo Zheng2, Shameen Afif-Rider1, Xiang Qing Yu2, Michele Gunsior2, Haihong Zhong3, Jay Harper3, Binyam Bezabeh4, Kapil Vashisht1, Marlon Rebelatto1, Molly Reed1, Patricia C. Ryan1, Shannon Breen3, Neki Patel5, Cui Chen3, Luke Masterson5, Arnaud Tiberghien5, Phillip W. Howard5, Nazzareno Dimasi4, and Rakesh Dixit1 Abstract Objective: To use preclinical models to identify a dosing gated to SG3249 (rats) or SG3400, a structurally related PBD schedule that improves tolerability of highly potent pyrroloben- (monkeys). zodiazepine dimers (PBDs) antibody drug conjugates (ADCs) Results: Observations of similar antitumor activity in mice without compromising antitumor activity. treated with single or fractionated dosing suggests that antitumor Experimental Design: A series of dose-fractionation studies activity of PBD ADCs is more closely related to total exposure were conducted to investigate the pharmacokinetic drivers (AUC) than peak drug concentrations (Cmax). In contrast, im- of safety and efficacy of PBD ADCs in animal models. The proved survival and reduced toxicity in rats and monkeys treated exposure–activity relationship was investigated in mouse with a fractionated dosing schedule suggests that tolerability of xenograft models of human prostate cancer, breast cancer, PBD ADCs is more closely associated with Cmax than AUC. and gastric cancer by comparing antitumor activity after Conclusions: We provide the first evidence that fractionated single and fractionated dosing with tumor-targeting ADCs dosing can improve preclinical tolerability of at least some conjugated to SG3249, a potent PBD dimer. The exposure– PBD ADCs without compromising efficacy. These findings tolerability relationship was similarly investigated in rat and suggest that preclinical exploration of dosing schedule could monkey toxicology studies by comparing tolerability, as be an important clinical strategy to improve the therapeutic assessed by survival, body weight, and organ-specifictoxi- window of highly potent ADCs and should be investigated cities, after single and fractionated dosing with ADCs conju- further. Clin Cancer Res; 1–11. Ó2017 AACR. Introduction the success of the two marketed products, clinical development of ADCs continues to be impeded by inability to escalate to Antibody drug conjugates (ADCs) are a rapidly growing class dosagerangeexpectedtoachievedesiredtherapeuticeffectsdue of targeted anticancer therapeutics that now account for a to safety issues (2). Of the greater than 50 ADCs evaluated in significant fraction of pharmaceutical pipelines (1). Much of early clinical trials, approximately 20 have been discontinued the interest in this technology stems from the recent marketing due to lack of efficacy and/or intolerable toxicity (3). Optimiz- approvals of ado-trastuzumab emtansine (Kadcyla) for the ing dosing schedule could be an important strategy to decrease treatment of metastatic breast cancer and brentuximab vedotin adverse events while maintaining exposure and efficacy (4). (Adcetris) for the treatment of Hodgkin's lymphoma. Despite Doing so requires a greater understanding of the relationship between exposure and toxicity in order to inform the clinical dosing schedule. The most common clinical dosing schedule 1Biologics Safety Assessment, MedImmune, Gaithersburg, Maryland. 2Clinical for ADCs is every 3 weeks (3). This schedule results in high peak 3 Pharmacology and DMPK, MedImmune, Gaithersburg, Maryland. Oncology plasma concentrations with minimal-to-no accumulation with Research, MedImmune, Gaithersburg, Maryland. 4Antibody Discovery and Pro- repeated dosing (5). Although every 3 weeks dosing is consis- tein Engineering, MedImmune, Gaithersburg, Maryland. 5Spirogen Ltd., QMB Innovation Centre, London, United Kingdom. tent with dosing schedules used for most chemotherapy regi- mens, it is unclear whether this represents the ideal dosing Note: Supplementary data for this article are available at Clinical Cancer Research Online (http://clincancerres.aacrjournals.org/). regimen for ADCs. In addition, there is very limited clinical data available to examine the potential benefits of alternative M.J.M. Hinrichs and P.M. Ryan contributed equally to this article. dosing schedules (3, 6, 7). SAR3419, an anti-CD19 ADC conju- Corresponding Author: Mary Jane Masson Hinrichs, MedImmune LLC, 1 Med- gated to the maytansinoid DM4, is an example where use of Immune Way, Gaithersburg, MD 20878. Phone: 2022957576; Fax: 3013981111; alternative dosing schedule positively impacted clinical devel- E-mail: [email protected] opment of an ADC. In the first in human trial, the maximum doi: 10.1158/1078-0432.CCR-17-0219 tolerated dose (MTD) was 160 mg/m2 on a every 3 weeks Ó2017 American Association for Cancer Research. dosing schedule (8). Although the overall response rate (ORR) www.aacrjournals.org OF1 Downloaded from clincancerres.aacrjournals.org on September 26, 2021. © 2017 American Association for Cancer Research. Published OnlineFirst June 19, 2017; DOI: 10.1158/1078-0432.CCR-17-0219 Hinrichs et al. risk of safety issues due to steep dose–response curves. Therefore, Translational Relevance understanding the relationship between exposure and tolerability Optimization of dosing schedule could be an important is essential to selecting a rational dosing schedule. We conducted a mitigation strategy to improve tolerability of ADCs with series of preclinical safety and efficacy studies to better understand narrow therapeutic windows, especially those conjugated to the relationship between exposure and therapeutic index of PBD highly potent payloads such as pyrrolobenzodiazepine dimers ADCs. Our aim was to identify whether dosing schedule can (PBDs). Doing so requires a greater understanding of the improve tolerability of highly potent ADC without compromising exposure–response relationship to enable selection of a dosing antitumor activity. regimen that decreases adverse events while maintaining effi- cacy. Assessing this relationship in patients can be lengthy and resource intensive; therefore, we evaluated whether preclinical Materials and Methods models could be used to understand the pharmacokinetic Test articles drivers of safety and efficacy for PBD ADCs in animals. By The anti-EphA2 (1C1), anti-5T4 (5T4), and isotype control conducting a series of dose-fractionation studies, we deter- (R347) antibodies used in these studies have been previously mined that fractionated dosing improved tolerability of PBD described (16–18). 1C1 cross-reacts with rat, monkey, and ADCs without impacting antitumor activity. Together, these human EphA2; 5T4 is only cross-reactive with monkey and results suggest that fractionated dosing could widen the ther- human 5T4 (16, 19). These antibodies were engineered to apeutic window of PBD ADCs. enable site-specific conjugation of two PBD dimers per anti- body. The maleimide-PEG8 PBD dimers used for conjugation were SG3249, also known as tesirine (20), and SG3400, a PBD dimer based on SG2000 (21, 22). Site-specific conjugation was carried out as described previously (17). Briefly, antibodies were at this dose level was 22%, findings of grade 2/3 ocular toxicity reduced using 40 molar equivalents of Tris (2-carboxy-ethyl)- prevented escalation to higher, potentially more efficacious phosphine (TCEP) in PBS pH 7.2, 1 mmol/L ethylenediamine dose levels (9). In an attempt to improve tolerability, a frac- tetraaceticacid (EDTA) for 3 hours at 37 C. Following overnight tionated dose schedule was evaluated in which patients were dialysis in PBS pH 7.2, 1 mmol/L EDTA at 4 C using 10,000 administered four weekly doses of 55 mg/m2 followed by four MWCO dialysis cassettes, 20 molar equivalents of dehydroas- additional doses every 2 weeks. Use of this schedule appeared corbic acid were added for 4 hours at 25 C. The solution was to have a positive impact as the ORR increased to 33%, and the filtered through a 0.2-mmsyringefilter and eight equivalents of severity of ocular toxicity decreased to mostly grade 1 events. SG3249 or SG3400 were sequentially added, followed by incu- Despite the success with SAR3419, reports with other ADCs bation at room temperature for 1 hour under gentle rotation. have been mixed. In the case of gemtuzumab ozogamicin The conjugation was quenched by the addition of 4 molar (Mylotarg), an anti-CD33 antibody conjugated to calicheami- equivalents (over SG3249 or SG3400) of N-acetyl cysteine. Free cin, early data from a multicenter phase II uncontrolled trial in unreacted SG3249 and SG3400 and macromolecular aggregates 57 adult patients with acute myeloblastic leukemia (AML), were removed using ceramic hydroxyapatite type II chromatog- patients demonstrated that fractionated dosing with mono- raphy as described previously (18). Site-specific ADCs were therapy Mylotarg significantly improved safety without impact- formulated at 3 mg/mL in PBS pH 7.2. The ADCs were char- ing efficacy (10). However, follow-up data from a small retro- acterized using complementary analytical methods and analyt- spective study in relapsed/refractory patients with AML treated ical data are shown in Supplementary Figure S1. with Mylotarg in combination with chemotherapy, using either the standard every 3 weeks (18 patients) or a fractionated (15 Animals